Identification of CC2D2A as a Meckel Syndrome Gene
Adds an Important Piece to the Ciliopathy Puzzle
Jonna Tallila,1Eveliina Jakkula,1,2Leena Peltonen,1,3,4,5Riitta Salonen,6and Marjo Kestila ¨1,*
Meckel syndrome (MKS) is a lethal malformation disorder characterized classically by encephalocele, polycystic kidneys, and polydac-
tyly. MKS is also one of the major contributors to syndromic neural tube defects (NTDs). Recent findings have shown primary cilia
dysfunction in the molecular background of MKS, indicating that cilia are critical for early human development. However, even though
four genes behind MKS have been identified to date, they elucidate only a minor proportion of the MKS cases. In this study, instead of
traditional linkage analysis, we selected 10 nonrelated affected fetuses and looked for the homozygous regions shared by them. Based on
this strategy, we identified the sixth locus and the fifth gene, CC2D2A (MKS6), behind MKS. The biological function of CC2D2A is
uncharacterized, but the corresponding polypeptide is predicted to be involved in ciliary functions and it has a calcium binding domain
(C2). Immunofluorescence staining of patient’s fibroblast cells demonstrates that the cells lack cilia, providing evidence for the critical
role of CC2D2A in cilia formation. Our finding is very significant not only to understand the molecular background of MKS, but also to
obtain additional information about the function of the cilia, which can help to understand their significance in normal development
and also in other ciliopathies, which are an increasing group of disorders with overlapping phenotypes.
Meckel syndrome (MKS [MIM 249000]) is an autosomal-
recessive lethal disorder characterized by a variety of severe
malformations. Minimal diagnostic criteria are cystic
dysplasia of the kidneys with fibrotic changes in the liver
and occipital encephalocele or some other central nervous
system malformation. Additionally, polydactyly is fre-
quently reported in the patients.1MKS diagnosis can be
established by ultrasound already in the end of the first
trimester.2Figures 1A and 1B show an ultrasound scan of
a MKS fetus at the 14thweek of gestation with encephalo-
cele and a distended stomach because of the enlarged
MKS is known to be a heterogenous disease with linkage
to five loci and four genes identified so far.3–7All the genes
are associated with ciliary functions. Mutations in ciliary
genes are known to cause a number of human monogenic
disorders that are collectively known as ciliopathies,
disorders with overlapping clinical features. An especially
interesting aspect of ciliary diseases is that they range
from embryonically lethal Meckel syndrome to less severe
multisystem disorders, such as Bardet-Biedl syndrome (BBS
[MIM 209900]), where the patients suffer from obesity, ret-
inal degeneration, polydactyly, mental retardation, and
cystic kidneys, for example.8
We have earlier reported that in 70% of the Finnish
cases, MKS is caused by the Finmajor(IVS15-7_35 del) muta-
tion in a novel MKS1 gene3that has later been identified
also in other populations.3,9–11In order to find out the ge-
netic defect behind MKS in the remaining Finnish families,
we carefully chose 10 out of 17 available fetuses that met
the minimal diagnostic criteria. This study has been ap-
proved by the ethical committees of the Joint Authority
for the Hospital District of Helsinki and Uusimaa, Finland.
Because these families had only one affected fetus and no
healthy siblings were available, a linkage-based position-
ing of the underlying locus was not possible. Instead,
assuming that in the isolated Finnish population the cases
might share a common mutation and surrounding haplo-
type, we decided to perform a genome-wide single-nucleo-
tide polymorphism (SNP) scan to locate homozygous re-
gions shared by the cases. DNA samples of MKS cases
were genotyped according to manufacturer’s instructions
on Illumina HumanHap300-duo SNP microarrays (Illu-
mina, San Diego, CA) containing 318,237 SNPs. All patient
samples had success rates of >99% and were thus included
in the study. Illumina Beadstudio v3.1.0 was used to call
genotypes, and homozygosity detector option was utilized
in the search of extended tracts of homozygosity in each
sample using a minimumlength of50 SNPs. The algorithm
gle SNP is homozygous in a sample, and it can be used to
autobookmark samples with extended tracts of homozy-
gosity (for more information, see Illumina Systems and
Software Technical Note).
Six out of ten patients were found to have overlapping
homozygous regions on chromosome 4p15. The size of
the homozygous regions varied from 730 kb to 6.8 Mb,
with all patients sharing a segment of 63 SNPs covering
a 565 kb area (Figure 2) (chr4: 14,909,996–15,475,912,
UCSC 2006). Illumina GenomeViewer was used to visual-
ize the chromosome 4 region to verify that the region
indeed was homozygous and to exclude copy-number
changes (data not shown). In addition, one patient shared
the same region, but was homozygote for a different allelic
haplotype. PLINK v1.0 was used to calculate pairwise iden-
tity-by-descent estimates and inbreeding coefficients.12
1National Public Health Institute and Institute for Molecular Medicine Finland, Helsinki 00290, Finland;2Institute for Molecular Medicine Finland, Finnish
Genome Center,3Department of Medical Genetics, University of Helsinki, Helsinki 00014, Finland;4The Broad Institute, Boston, MA 02142, USA;5The
Wellcome Trust Sanger Institute, Hinxton CB10 1SA, UK;6Department of Medical Genetics, Va ¨esto ¨liitto, Helsinki 00100, Finland
DOI 10.1016/j.ajhg.2008.05.004. ª2008 by The American Society of Human Genetics. All rights reserved.
The American Journal of Human Genetics 82, 1361–1367, June 2008
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The American Journal of Human Genetics 82, 1361–1367, June 2008